Related Experiment Video
Updated: Aug 3, 2025
Pooled shRNA Library Screening to Identify Factors that Modulate a Drug Resistance Phenotype
Published on: June 17, 2022
Interrogating bromodomain inhibitor resistance in KMT2A-rearranged leukemia through combinatorial CRISPR screens
Shaela Wright1, Jianzhong Hu2, Hong Wang3
1Department of Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, TN 38105.
Abstract:
Bromo- and extra-terminal domain inhibitors (BETi) have exhibited therapeutic activities in many cancers. However, the mechanisms controlling BETi response and resistance are not well understood. We conducted genome-wide loss-of-function CRISPR screens using BETi-treated KMT2A-rearranged (KMT2A-r) cell lines. We revealed that Speckle-type POZ protein (SPOP) gene (Speckle Type BTB/POZ Protein) deficiency caused significant BETi resistance, which was further validated in cell lines and xenograft models. Proteomics analysis and a kinase-vulnerability CRISPR screen indicated that cells treated with BETi are sensitive to GSK3 perturbation. Pharmaceutical inhibition of GSK3 reversed the BETi-resistance phenotype. Based on this observation, a combination therapy regimen inhibiting both BET and GSK3 was developed to impede KMT2A-r leukemia progression in patient-derived xenografts in vivo. Our results revealed molecular mechanisms underlying BETi resistance and a promising combination treatment regimen of ABBV-744 and CHIR-98014 by utilizing unique ex vivo and in vivo KMT2A-r PDX models.
Insights
Speckle-type POZ protein (SPOP) deficiency drives resistance to Bromo- and extra-terminal domain inhibitors (BETi) in KMT2A-rearranged leukemia. Combining BET inhibitors with GSK3 inhibitors overcomes this resistance, offering a new therapeutic strategy.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Bromo- and extra-terminal domain inhibitors (BETi) show promise in cancer therapy.
- Mechanisms of BETi response and resistance, particularly in KMT2A-rearranged (KMT2A-r) leukemia, remain unclear.
Purpose of the Study:
- To elucidate mechanisms of BETi resistance in KMT2A-r cancer.
- To identify novel therapeutic strategies for overcoming BETi resistance.
Main Methods:
- Genome-wide CRISPR screens in BETi-treated KMT2A-r cell lines.
- Validation in cell lines and xenograft models.
- Proteomics analysis and kinase-vulnerability screens.
- Combination therapy studies in patient-derived xenografts (PDX).
Main Results:
- Deficiency in Speckle-type POZ protein (SPOP) confers significant BETi resistance.
- BETi-treated cells are vulnerable to Glycogen Synthase Kinase 3 (GSK3) inhibition.
- Pharmacological GSK3 inhibition reverses BETi resistance.
- A combination of BETi (ABBV-744) and GSK3 inhibitor (CHIR-98014) effectively impedes KMT2A-r leukemia progression in vivo.
Conclusions:
- SPOP deficiency is a key mechanism of BETi resistance.
- Targeting GSK3 in combination with BETi presents a promising therapeutic approach for KMT2A-r leukemia.
